Understanding the reasons why SIV-infected sooty mangabeys (SMs) remain healthy despite high viremia is a key unanswered question in contemporary AIDS research, with important ramifications in terms of HIV pathogenesis, therapy, and vaccines. In recent studies, we have sorted central memory CD4+ T cells (TCM) and effector-memory CD4+ T cells (CD4+ TEM) from SIV-infected SMs and rhesus macaques (RMs), and that, while CD4+ TEM were similarly infected in both species, CD4+ TCM of SMs show significantly (>1 log) fewer SIV-DNA copies in vivo than CD4+ TCM of RMs. Based on this result, we hypothesize that protection of CD4+ TCM from virus infection is a key mechanisms by which SIV-infected SMs avoid CD4+ T cell depletion, chronic immune activation, and progression to AIDS. To test this hypothesis we will expand upon this previous work and propose a series of studies that will clarify the main features of in vivo and in vitro SIV infection in CD4+ TCM of both SMs and RMs, and elucidate the mechanisms by which CD4+ TCM of SMs are protected from SIV infection. In the first Aim, we will conduct a systematic comparative analysis of in vivo SIV replication in different CD4+ T cell subsets of SIV-infected SMs and RMs. We will examine different tissues (blood, lymph nodes, mucosal tissues) and CD4+ T cell subsets (Th1, Th2, Th17, and Tregs), and establish correlations with the course of disease progression. In the second Aim, we will study the relationship between SIV replication in central memory CD4+ T cells of LNs and the presence of LN immunopathology, including loss of CD4+ T cells, immune activation, disruption of architecture, fibrosis, and follicular DC virus trapping. In the third Aim, we will assess how SIV replicates in vivo in CD4+ TCM and CD4+ TEM of SMs and RMs by (i) conducting a detailed analysis of the level of integrated proviral DNA, the frequency of SIV-infected cells and the number of RNA copies/cell by limiting dilution analysis, and by (ii) performing sequence analysis of the virus. In the fourth Aim, we will assess the intrinsic susceptibility to SIV infection by CD4+ TCM and CD4+ TEM of uninfected SMs and RMs by conducting experiments of in vitro infection, in which we will seek to identify what stage(s) of virus replication are blocked in CD4+ TCM of SMs. As part of this Aim, we will also study the expression of host restriction factors (i.e., TRIM-5a, APOBECs, and Tetherin) as well as the global transcriptional profile in CD4+ TCM and CD4+ TEM of SMs and RMs. We believe that these studies will advance significantly our understanding of how naturally SIV-infected SMs are resistant to AIDS despite high viremia. We envision that answering this question will provide clues to AIDS pathogenesis in humans that will have ultimately an impact on the prevention and treatment of HIV infection.